Conjugal transfer of aac(6')Ie-aph(2″)Ia gene from native species and mechanism of regulation and cross resistance in Enterococcus faecalis MCC3063 by real time-PCR.

High level aminoglycoside resistance (HLAR) in the lactic acid bacteria (LAB) derived from food animals is detrimental. The aim of this study was to investigate the localization and conjugal transfer of aminoglycoside resistance genes, aac(6')Ie-aph(2″)Ia and aph(3')IIIa in different Enterococcus species. The cross resistance patterns in Enterococcus faecalis MCC3063 to clinically important aminoglycosides by real time PCR were also studied. Southern hybridization experiments revealed the presence of aac(6')Ie-aph(2″)Ia and aph(3')IIIa genes conferring HLAR in high molecular weight plasmids except in Lactobacillus plantarum. The plasmid encoded bifunctional aac(6')Ie-aph(2″)Ia gene was transferable from Enterococcus avium (n = 2), E. cecorum (n = 1), E. faecalis (n = 1) and Pediococcus lolii (n = 1) species into the recipient strain; E. faecalis JH2-2 by filter mating experiments thus indicating the possible risks of gene transfer into pathogenic strains. Molecular analysis of cross resistance patterns in native isolate of E. faecalis MCC3063 carrying aac(6')Ie-aph(2″)Ia and aph(3')IIIa gene was displayed by quantification of the mRNA levels in this study. For this, the culture was induced with increasing concentrations of gentamicin, kanamycin and streptomycin (2048, 4096, 8192, 16384 µg/mL) individually. The increasing concentrations of gentamicin and kanamycin induced the expression of the aac(6')Ie-aph(2″)Ia and aph(3')IIIa resistance genes, respectively. Interestingly, it was observed that induction with streptomycin triggered a significant fold increase in the expression of the aph(3')IIIa gene which otherwise was not known to modify the aminoglycoside. This is noteworthy as streptomycin was found to confer cross resistance to structurally unrelated kanamycin. Also, expression of the aph(3')IIIa gene when induced with streptomycin, revealed that bacteria harbouring this gene will be able to overcome streptomycin bactericidal action at specific concentrations. HLAR in E. faecalis MCC3063 may be due to the combined expression of both the aac(6')Ie-aph(2″)Ia and aph(3')IIIa genes which could be therapeutically challenging. A combined expression of both the genes in E. faecalis MCC3063 may yield HLAR which could be therapeutically challenging. The study highlights the significant alterations in the mRNA expression levels of aac(6')Ie-aph(2″)Ia and aph(3')IIIa in resistant pathogens, upon exposure to clinically vital aminoglycosides.

Assessment of probiotic, antifungal and cholesterol lowering properties of Pediococcus pentosaceus KCC-23 isolated from Italian ryegrass.

BACKGROUND: Lactic acid bacteria (LAB) are important for the processing of various food products. Although genetically modified organisms have contributed to improvements in various food products, there are some limitations. Thus, the discovery of wild strains from natural sources must be considered as the most suitable approach for identifying new LAB. Therefore, we planned to isolate and characterise the LAB from Italian ryegrass forage and evaluate their biological potential. RESULTS: A total of 28 strains were isolated and screened for their anti-fungal and probiotic properties. A single strain was selected due to its antifungal and probiotic efficiency. The strain was identified as Pediococcus pentosaceus KCC-23. The strain KCC-23 showed effective inhibition against Aspergillus fumigatus, Pencillium chrysogenum, Pencillium roqueforti, Botrytis elliptica and Fusarium oxysporum. Further, it survived low pH, and the presence of bile salts and gastric juice. It exhibited significant aggregation and hydrophobicity properties. The KCC-23 effectively assimilated cholesterol and had the ability to utilise pre-biotics such as raffinose and inulin. Finally, KCC-23 exhibited significant free radical scavenging activity. CONCLUSION: P. pentosaceus KCC-23 showed effective anti-fungal, probiotic and anti-oxidant properties and would be a promising isolate for exploitation in the formulation of food for ruminants and humans.

Analysis of cell growth dynamics of Pediococcus acidilactici in the presence of inulin in an optimized microenvironment.

The present investigation deals with the optimization of cell growth rate of the candidate probiotic Pediococcus acidilactici in the presence of the specific prebiotic inulin. Three independent variables viz. concentration of inulin, concentration of glucose and pH have been selected for optimization study using response surface methodology. Theoretical analysis indicates that the maximum cell growth rate occurs at pH 7, 20 g/dm(3) concentration of inulin and 20 g/dm(3) concentration of glucose. Validation of these values has been done through a set of programmed experiments. Studies on cell dynamics in the presence of different concentrations of inulin have also been carried out to identify any limitation on the initial inulin concentration. Results clearly indicate that cell growth is enhanced with the increase in inulin concentration. However, there is a critical value of the prebiotic concentration (20 g/dm(3) inulin) beyond which the cell growth is inhibited. A summative type growth model has been proposed to explain the growth behaviour of P. acidilactici in the presence of the dual substrate, i.e. glucose and inulin. While growth on glucose follows Monod model, Haldane-type substrate-inhibited growth model holds good for growth on inulin. Intrinsic kinetic parameters for all the model equations have been determined experimentally.

Characterization of the phenolic fraction from Argentine wine and its effect on viability and polysaccharide production of Pediococcus pentosaceus.

OBJECTIVES: To qualitatively and quantitatively characterize a low molecular weight phenolic fraction (LMF) of Malbec wine from Cafayate, Argentina, and evaluate its effect on viability and exopolysaccharide production of Pediococcus pentosaceus 12p, a wine spoilage bacterium. RESULTS: The phenolic compounds detected were, in general, comparable to data previously reported but hydroxycinnamic acids were detected at higher concentrations than determined in other studies. Addition of LMF at identical concentrations present in wine or a four times concentrated LMF mixture to a synthetic wine-like medium produced a diminution in bacterial viability and exopolysaccharide production in the supernatant culture. Transmission electron microscopy revealed damage of bacterial cell integrity after 96 h of incubation only in the presence of four times concentrated LMF. CONCLUSION: This is the first time a low molecular weight phenolic fraction has been characterized in Cafayate wine and it has demonstrated a marked antimicrobial effect on an exopolysaccharide-producing wine spoilage bacterium.

Short communication: 3-phenyllactic acid production in milk by Pediococcus pentosaceus SK25 during laboratory fermentation process.

3-Phenyllactic acid (PLA) is a broad-spectrum antimicrobial compound, produced by a wide range of lactic acid bacteria. A novel lactic acid bacteria strain with high PLA-producing ability, Pediococcus pentosaceus SK25, was isolated from traditional Chinese pickles. When grown in de Man, Rogosa, Sharpe broth at 30°C for 36h, this strain produced 135.6mg/L of PLA. Using this strain as starter for milk fermentation, 47.2mg/L of PLA was produced after fermentation for 12h. The PLA production was significantly improved by phenylalanine supplement, but was completely inhibited by tyrosine supplement.

In situ production of human ß defensin-3 in lager yeasts provides bactericidal activity against beer-spoiling bacteria under fermentation conditions.

AIMS: To examine the use of a natural antimicrobial peptide, human ß-defensin-3 (HBD3), as a means of preventing spoilage from bacterial contamination in brewery fermentations and in bottled beer. METHODS AND RESULTS: A chemically synthesised HBD3 peptide was tested for bactericidal activity against common Gram-positive and Gram-negative beer-spoiling bacteria, including species of Lactobacillus, Pediococcus and Pectinatus. The peptide was effective at the µmol l(-1) range in vitro, reducing bacterial counts by 95%. A gene construct encoding a secretable form of HBD3 was integrated into the genome of the lager yeast Saccharomyces pastorianus strain CMBS-33. The integrated gene was expressed under fermentation conditions and was secreted from the cell into the medium, but a significant amount remains associated with yeast cell surface. We demonstrate that under pilot-scale fermentation conditions, secreted HBD3 possesses bactericidal activity against beer-spoiling bacteria. Furthermore, when added to bottled beer, a synthetic form of HBD3 reduces the growth of beer-spoiling bacteria. CONCLUSIONS: Defensins provide prophylactic protection against beer-spoiling bacteria under brewing conditions and also in bottled beer. SIGNIFICANCE AND IMPACT OF THE STUDY: The results have direct application to the brewing industry where beer spoilage due to bacterial contamination continues to be a major problem in breweries around the world.

Probiotic properties of Pediococcus strains isolated from jeotgals, salted and fermented Korean sea-food.

Three Pediococcus pentosaceus strains were isolated from jeotgals, salted and fermented Korean sea-foods, and their probiotic potentials were examined. After 2 h exposure to pH 3.0, P. pentosaceus F66 survived with the survival ratio of 32.6% followed by P. pentosaceus D56 (17.2%) and P. pentosaceus A24 (7.5%). P. pentosaceus F66 also survived better (26.6%) than P. pentosaceus A24 (13.7%) and P. pentosaceus D56 (5.8%) after 2 h exposure to 0.3% bile salts. Three strains grew slowly on MRS broth with 15% NaCl (w/v), reaching the OD600 values of 0.4-0.8 in 36 h. They adhered to Caco-2 cells (10.9-13.9 CFU/cell) with similar degree of adherence of a positive control, Lactobacillus rhamnosus GG (12.8 ± 0.5 CFU/cell). Three strains possess some desirable enzyme activities such as ß-galactosidase, α-glucosidase, ß-glucosidase, and N-acetyl-ß-glucosidase. From these results, P. pentosaceus F66 seems qualified as a probiotic and can be utilized for fermented foods including jeotgals.

Yerba mate enhances probiotic bacteria growth in vitro but as a feed additive does not reduce Salmonella Enteritidis colonization in vivo.

Yerba mate (Ilex paraguariensis) is a tea known to have beneficial effects on human health and antimicrobial activity against some foodborne pathogens. Thus, the application of yerba mate as a feed additive for broiler chickens to reduce Salmonella colonization was evaluated. The first in vitro evaluation was conducted by suspending Salmonella Enteritidis and lactic acid bacteria (LAB) in yerba mate extract. The in vivo evaluations were conducted using preventative and horizontal transmission experiments. In all experiments, day-of-hatch chicks were treated with one of the following 1) no treatment (control); 2) ground yerba mate in feed; 3) probiotic treatment (Lactobacillus acidophilus and Pediococcus; 9:1 administered once on day of hatch by gavage); or 4) both yerba mate and probiotic treatments. At d 3, all chicks were challenged with Salmonella Enteritidis (preventative experiment) or 5 of 20 chicks (horizontal transmission experiment). At d 10, all birds were euthanized, weighed, and cecal contents enumerated for Salmonella. For the in vitro evaluation, antimicrobial activity was observed against Salmonella and the same treatment enhanced growth of LAB. For in vivo evaluations, none of the yerba mate treatments significantly reduced Salmonella Enteritidis colonization, whereas the probiotic treatment significantly reduced Salmonella colonization in the horizontal transmission experiment. Yerba mate decreased chicken BW and decreased the performance of the probiotic treatment when used in combination. In conclusion, yerba mate had antimicrobial activity against foodborne pathogens and enhanced the growth of LAB in vitro, but in vivo yerba mate did not decrease Salmonella Enteritidis colonization.

Lethal hydroxyl radical accumulation by a lactococcal bacteriocin, lacticin Q.

The antimicrobial mechanism of a lactococcal bacteriocin, lacticin Q, can be described by the toroidal pore model without any receptor. However, lacticin Q showed different degrees of activity (selective antimicrobial activity) against Gram-positive bacteria even among related species. The ability of lacticin Q to induce pore formation in liposomes composed of lipids from different indicator strains indicated that its selective antimicrobial activity could not be attributed only to membrane lipid composition. We investigated the accumulation of deleterious hydroxyl radicals after exposure to lacticin Q as a contributing factor to cell death in the indicator strains. When lacticin Q of the same concentration as the MIC or minimum bactericidal concentration was added to the indicator cultures, high levels of hydroxyl radical accumulation were detected. Treatment with hydroxyl radical scavengers, thiourea and 2,2'-bipyridyl, decreased the levels of hydroxyl radical accumulation and recovered cell viability. These results suggest that, with or without pore formation, the final antimicrobial mechanism of lacticin Q is the accumulation of hydroxyl radicals, which varies by strain, resulting in the selective antimicrobial activity of lacticin Q.

Determination of the mode of action of enterolysin A, produced by Enterococcus faecalis B9510.

AIM: The current study aimed to visualize the damage caused by enterolysin A to the cells of sensitive strains and to find out cleavage site within the peptidoglycan moiety of bacterial cell walls. METHODS AND RESULTS: Enterolysin A produced by a local isolate, Enterococcus faecalis B9510 was found to rapidly kill cells of the sensitive strain Lactococcus lactis ssp. cremoris 2144 during 120 min of treatment as compared to the untreated control where no such effect was observed. Transmission electron microscopy of the enterolysin A-treated cells revealed leaking of the cytoplasmic contents ultimately resulting in complete lysis of cell walls. To find the cleavage site, purified cell walls of L. lactis ssp. cremoris 2144, Pediococcus pentosaceus 43201 and Lactobacillus delbrueckii ssp. bulgaricus ATCC 11842 were treated with enterolysin A, and liberated amino acids were derivatized for N and C terminals and analysed using thin layer chromatography on silica gel with isopropanol as solvent. The results showed that enterolysin A cleaves the peptide bonds at two locations within peptidoglycan subunits. The first location is between L-alanine and D-glutamic acid of the stem peptide and the other location is between L-lysine of the stem peptide and D-aspartic acid of the interpeptide bridge. CONCLUSIONS: Enterolysin A cleaves the peptide bonds within the stem peptide as well as in the interpeptide bridge of Gram-positive bacterial cell walls. This gives a possible reason for the broad spectrum of enterolysin A activity. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report identifying the cleavage site of enterolysin A within the cell walls of sensitive bacteria. This will help in identifying potential applications for enterolysin A.

BlpC-regulated bacteriocin production in Streptococcus thermophilus.

Streptococcus thermophilus B59671 produces a bacteriocin with anti-pediococcal activity, but genes required for its production are not characterized. Genome sequencing of S. thermophilus has identified a genetic locus encoding a quorum sensing (QS) system that regulates production of class II bacteriocins. However, in strains possessing this gene cluster, production of bacteriocin like peptides (Blp) was only observed when excess pheromone was provided. PCR analysis revealed this strain possessed blpC, which encodes the 30-mer QS pheromone. To investigate if BlpC regulates bacteriocin production in S. thermophilus B59671, an integrative vector was used to replace blpC with a gene encoding for kanamycin resistance and the resulting mutant did not inhibit the growth of Pediococcus acidilactici. Constitutive expression of blpC from a shuttle vector restored the bacteriocin production, confirming the blp gene cluster is essential for bacteriocin activity in S. thermophilus B59671.

The synthesis of active and stable diaminopimelate analogues of the lantibiotic peptide lactocin S.

Lantibiotic peptides are potent antimicrobial compounds produced by Gram-positive bacteria. They can be used in food preservation, and some also show potential for clinical applications. Unfortunately, some of these peptides can be susceptible to inactivation by oxidation of the sulfur-containing amino acid lanthionine, limiting their use. Here we describe the synthesis and testing of diaminopimelate analogues of the lantibiotic lactocin S. These analogues were designed to improve the oxidative stability of the peptide by replacing the sulfur in lanthionine with a methylene unit. Lanthionine was systematically replaced with diaminopimelate during solid-phase peptide synthesis to produce several analogues. One analogue, A-DAP lactocin S, was found to retain full biological activity in addition to displaying increased stability. This is the first time a synthetic lanthionine ring analogue of a lantibiotic has retained natural activity levels. This methodology is potentially very promising for use in producing more stable, medically relevant lantibiotics.

Development of a novel probiotic delivery system based on microencapsulation with protectants.

The establishment of the health-promoting benefits of probiotics is challenged by the antimicrobial bio-barriers throughout the host's gastrointestinal (GI) tract after oral administration. Although microencapsulation has been frequently utilised to enhance the delivery of probiotics, microcapsules of sub-100 µm were found to be ineffective and therefore questioned as an effective delivery vehicle for viable probiotics despite the sensory advantage. In this study, four probiotics strains were encapsulated in chitosan-coated alginate microcapsules of sub-100 µm. Only a minor protective effect was observed from this original type of microcapsule. In order to enhance the survival of these probiotics, sucrose, a metabolisable sugar, and lecithin vesicles were added to the wall material. Both of the ingredients could be readily encapsulated with the probiotics, and protected them from stresses in the simulated GI fluids. The metabolisable sugar effectively increased the survival of the probiotics in gastric acid, mainly through energizing the membrane-bound F1F0-ATPases. The lecithin vesicles proved to alleviate the bile salt stress, and hence notably reduced the viability loss at the elevated bile salt concentrations. Overall, three out of the total four probiotics in the reinforced sub-100 µm microencapsules could significantly survive through an 8-h sequential treatment of the simulated GI fluids, giving less than 1-log drop in viable count. The most vulnerable strain of bifidobacteria also yielded a viability increase of 3-logs from this protection. In conclusion, the sub-100 µm microcapsules can be a useful vehicle for the delivery of probiotics, as long as suitable protectants are incorporated in the wall matrix.

Wine phenolic compounds influence the production of volatile phenols by wine-related lactic acid bacteria.

AIMS: To evaluate the effect of wine phenolic compounds on the production of volatile phenols (4-vinylphenol [4VP] and 4-ethylphenol [4EP]) from the metabolism of p-coumaric acid by lactic acid bacteria (LAB). METHODS AND RESULTS: Lactobacillus plantarum, Lactobacillus collinoides and Pediococcus pentosaceus were grown in MRS medium supplemented with p-coumaric acid, in the presence of different phenolic compounds: nonflavonoids (hydroxycinnamic and benzoic acids) and flavonoids (flavonols and flavanols). The inducibility of the enzymes involved in the p-coumaric acid metabolism was studied in resting cells. The hydroxycinnamic acids tested stimulated the capacity of LAB to synthesize volatile phenols. Growth in the presence of hydroxycinnamic acids, especially caffeic acid, induced the production of 4VP by resting cells. The hydroxybenzoic acids did not significantly affect the behaviour of the studied strains. Some of the flavonoids showed an effect on the production of volatile phenols, although strongly dependent on the bacterial species. Relatively high concentrations (1 g l(-1) ) of tannins inhibited the synthesis of 4VP by Lact. plantarum. CONCLUSIONS: Hydroxycinnamic acids were the main compounds stimulating the production of volatile phenols by LAB. The results suggest that caffeic and ferulic acids induce the synthesis of the cinnamate decarboxylase involved in the metabolism of p-coumaric acid. On the other hand, tannins exert an inhibitory effect. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights the capacity of LAB to produce volatile phenols and that this activity is markedly influenced by the phenolic composition of the medium.

Potential beneficial properties of bacteriocin-producing lactic acid bacteria isolated from smoked salmon.

AIMS: To evaluate the probiotic properties of strains isolated from smoked salmon and previously identified as bacteriocin producers. METHODS AND RESULTS: Strains Lactobacillus curvatus ET06, ET30 and ET31, Lactobacillus fermentum ET35, Lactobacillus delbrueckii ET32, Pediococcus acidilactici ET34 and Enterococcus faecium ET05, ET12 and ET88 survived conditions simulating the gastrointestinal tract (GIT) and produced bacteriocins active against several strains of Listeria monocytogenes, but presented very low activity against other lactic acid bacteria (LAB). Cell-free supernatants containing bacteriocins, added to 3-h-old cultures of L. monocytogenes 603, suppressed growth over 12 h. Auto-aggregation was strain-specific, and values ranged from 7·2% for ET35 to 12·1% for ET05. Various degrees of co-aggregation with L. monocytogenes 603, Lactobacillus sakei ATCC 15521 and Enterococcus faecalis ATCC 19443 were observed. Adherence of the bacteriocinogenic strains to Caco-2 cells was within the range reported for Lactobacillus rhamnosus GG, a well-known probiotic. The highest levels of hydrophobicity were recorded for Lact. curvatus (61·9­64·6%), Lact. fermentum (78·9%), Lact. delbrueckii (43·7%) and Ped. acidilactici (51·3%), which are higher than the one recorded for Lact. rhamnosus GG (53·3%). These strains were highly sensitive to several antibiotics and affected by several drugs from different generic groups in a strain-dependent manner. CONCLUSIONS: Smoked salmon is a rich source of probiotic LAB. All strains survived conditions simulating the GIT and produced bacteriocins active against various pathogens. Adherence to Caco-2 cells was within the range reported for Lact. rhamnosus GG, a well-known probiotic. In addition, the high hydrophobicity readings recorded define the strains as good probiotics.

Use of the yeast Pichia pastoris as an expression host for secretion of enterocin L50, a leaderless two-peptide (L50A and L50B) bacteriocin from Enterococcus faecium L50.

In this work, we report the expression and secretion of the leaderless two-peptide (EntL50A and EntL50B) bacteriocin enterocin L50 from Enterococcus faecium L50 by the methylotrophic yeast Pichia pastoris X-33. The bacteriocin structural genes entL50A and entL50B were fused to the Saccharomyces cerevisiae gene region encoding the mating pheromone alpha-factor 1 secretion signal (MFalpha1(s)) and cloned, separately and together (entL50AB), into the P. pastoris expression and secretion vector pPICZalphaA, which contains the methanol-inducible alcohol oxidase promoter (P(AOX1)) to express the fusion genes. After transfer into the yeast, the recombinant plasmids were integrated into the genome, resulting in three bacteriocinogenic yeast strains able to produce and secrete the individual bacteriocin peptides EntL50A and EntL50B separately and together. The secretion was efficiently directed by MFalpha1(s) through the Sec system, and the precursor peptides were found to be correctly processed to form mature and active bacteriocin peptides. The present work describes for the first time the heterologous expression and secretion of a two-peptide non-pediocin-like bacteriocin by a yeast.

Susceptibility of meat starter cultures to antimicrobials used in food animals in Canada.

Lactic acid bacteria (LAB) are extensively used in the food industry for fermentation processes. However, it is possible that these bacteria may serve as a reservoir for antibiotic resistance genes that can be transferred to pathogens, giving rise to public health concerns. Animal operations that use antimicrobials as growth promotants have been linked to the origin of resistance due to the selective effect of low levels of antimicrobial used in this management strategy. The objective of this study was to determine the antimicrobial susceptibilities and mechanisms of resistance for 30 isolates of meat starter cultures commonly used in dry sausage fermentations to 20 antimicrobial agents. Susceptibility tests were performed by broth microdilution using Iso-Sensitest broth (90%, vol/vol) and de Man Rogosa Sharpe (MRS) broth (10%, vol/vol). The results showed that all 30 isolates exhibited resistance to at least three antimicrobials regardless of antimicrobial class while 17 or 30% of strains were resistant to antibiotics in three or six different classes, respectively. The incidence of antimicrobial resistance was higher among Pediococcus pentosaceus and lower for Staphylococcus carnosus strains. Genetic determinants for the lincosamide, macrolide, and tetracycline antimicrobials were not found using PCR. Phenotypic resistance in the absence of known resistance genes found here suggests that other mechanisms or genes might have contributed to the negative results. Further studies are needed to explore the genetic mechanisms underlying the prevalence of antibiotic resistance in Pediococcus species.

Increased inactivation of exopolysaccharide-producing Pediococcus parvulus in apple juice by combined treatment with enterocin AS-48 and high-intensity pulsed electric field.

The cyclic peptide bacteriocin enterocin AS-48 was tested (at final concentrations of 0.175, 0.613, and 1.05 AU/ml) against the exopolysaccharide-producing cider spoilage strain Pediococcus parvulus 48 in apple juice in combination with high-intensity pulsed electric field (HIPEF) treatment (35 kV/cm and 150 Hz for 4 mus and bipolar mode). The effect of the combined treatments was studied by surface response methodology, with AS-48 concentration and HIPEF treatment time as process variables. A bacteriocin concentration of 0.613 AU/ml in combination with HIPEF treatment time of 1,000 micros reduced the population of pediococci by 6.6 log cycles in apple juice and yielded an apple juice that was free from pediococci during a 30-day storage period at 4 and 22 degrees Celsius. In contrast, application of HIPEF treatment alone had no effect on the surviving pediococci during storage of juice at 22 degrees Celsius. The combined treatment significantly improved the stability of the juice against spoilage by exopolysaccharide-producing P. parvulus.

Assessment of potential probiotic- and starter properties of Pediococcus spp. isolated from Turkish-type fermented sausages (sucuk).

In this study, the metabolic activities of five strains of amount of Pediococcus spp. in terms of the quantities they produced of lactic acid, hydrogen peroxide, exopolysaccharides and proteolytic activity were determined. Lactic acid levels produced by these strains were found to be in the range of 2.5-5.6 mg/ml. All strains produced hydrogen peroxide. The P. pentosaceus Z13P strain produced the maximum amount (0.25 mg/ml) of proteolytic activity. Exopolysaccharide (EPS) production by the Pediococcus strains during growth in MRS medium was in the range 25-64 mg/l. The susceptibility of 10 different antibiotics against these strains was also tested. All strains were found to be resistant to amoxicillin, gentamicin, and vancomicin. Antimicrobial effects of the Pediococcus on pathogens were also determined by an agar diffusion method. All of the strains were able to inhibit L. monocytogenes. The tolerance of the strains to low pH, their resistance to bile salts of strains, and their abilities to autoaggregate and coaggregate with L. monocytogenes were also evaluated.

Susceptibility of Pediococcus isolates to antimicrobial compounds in relation to hop-resistance and beer-spoilage.

BACKGROUND: Though important in the context of food microbiology and as potential pathogens in immuno-compromised humans, bacterial isolates belonging to the genus Pediococcus are best known for their association with contamination of ethanol fermentation processes (beer, wine, or fuel ethanol). Use of antimicrobial compounds (e.g., hop-compounds, Penicillin) by some industries to combat Pediococcus contaminants is long-standing, yet knowledge about the resistance of pediococci to antimicrobial agents is minimal. Here we examined Pediococcus isolates to determine whether antibiotic resistance is associated with resistance to hops, presence of genes known to correlate with beer spoilage, or with ability to grow in beer. RESULTS: Lactic acid bacteria susceptibility test broth medium (LSM) used in combination with commercially available GPN3F antimicrobial susceptibility plates was an effective method for assessing antimicrobial susceptibility of Pediococcus isolates. We report the finding of Vancomycin-susceptible Pediococcus isolates from four species. Interestingly, we found that hop-resistant, beer-spoilage, and beer-spoilage gene-harbouring isolates had a tendency to be more susceptible, rather than more resistant, to antimicrobial compounds. CONCLUSION: Our findings indicate that the mechanisms involved in conferring hop-resistance or ability to spoil beer by Pediococcus isolates are not associated with resistance to antibiotics commonly used for treatment of human infections. Also, Vancomycin-resistance was found to be isolate-specific and not intrinsic to the genus as previously believed.